REGULATION OF INTEGRIN-MEDIATED ADHESION BY MUSCARINIC ACETYLCHOLINE-RECEPTORS AND PROTEIN-KINASE-C IN SMALL-CELL LUNG-CARCINOMA

Study objectives: Improved understanding of the phenotypic characteris tics of small cell lung cancer (SCLC) cells may facilitate the develop ment of new therapies for this bronchogenic malignancy with early meta stases. Herein we investigate whether activation of the M-3 subtype of muscarinic acetylcholine receptor (m-AChR) expressed on SCLC cells af fects beta(1)-integrin-mediated adhesion of these cells. Design: Adhes ion of the SCLC cell lines SCC-9 and NCI-H345 to extracellular matrix (ECM) proteins was investigated. Cell adhesion was quantified by label ing the cells with either toluidine blue dye and measuring optical den sity or H-3-thymidine and measuring beta-activity. Fluorescence-activa ted cell sorting was used to quantify the SCLC cell surface expression of beta(1)-integrins. Setting: Experiments were conducted in the Mole cular Pharmacology Laboratory, Guthrie Research Institute. Measurement s and results: Activation of mAChR with the agonist carbachol (10 mu M , 1.5 h) significantly increases adhesion of die SCC-9 SCLC cell line to the ECM proteins laminin and collagen types I and IV. In contrast, mAChR activation does not alter the adhesion of SCC-9 cells to vitrone ctin, fibronectin, poly-L-lysine, or bovine serum albumin, Carbachol a lso does not alter the adhesion of NCI-H345 SCLC cells that Lack funct ional mAChR. Preincubation of SCC-9 cells with the AIIB2 blocking anti body to beta(1)-integrin inhibits mAChR-induced adhesion to ECM protei ns, Immunofluorescence analysis indicates that mAChR activation does n ot alter the surface expression of beta(1)-integrins by SCC-9 cells. D irect stimulation of protein kinase C (PKC) by treatment with phorbol 12-myristate 13-acetate (PMA) (10 nM, 1.5 h) increases the adhesion of both the SCC-9 and NCI-H345 cell lines to ECM proteins. These results indicate that direct activation of PRC or stimulation of M-3 mAChR (w hich results in increased PKC activity) increases the binding activity of beta(1)-integrins, resulting in increased adhesion of SCLC cells t o ECM proteins. Conclusions: The ability of mAChR to regulate SCLC pro liferation and adhesion suggests that activation of these receptors ma y be used to alter SCLC tumorigenesis and metastasis.